This project is directed, broadly, at developing and using physical and mathematical methodologies to understand integrative aspects of complex cellular processes. Emphasis has been on studying the functioning of multicomponent, supramolecular entities such as composite membranes and elements of cytoskeletal networks, particularly with a view towards understanding their involvement in cellular organization and cell shape. Cognate in vitro reconstitution studies of purified cell components also have been undertaken. These investigations oftentimes employ unusual physical instrumentation, in part to examine the relevance and utility of such devices for biological research. For example, a fluorescence correlation microscope (FCM) recently has been constructed which now is being used to investigate the sizes and stability of aberrant tubulin ring polymers induced by dolastatin, cryptophysin, and similar antimitotic agents. This study is of intrinsic interest, as these ring-forming compounds currently are being evaluated elsewhere as possible anti-tumor drugs, but it also serves to calibrate the instrument and drive development of new theory for interpreting measurements. Similar studies of other multi-component cellular assemblies are planned, among which are investigations of the size distributions of reconstituted clathrin baskets. In this case, measurements will be related to molecular parameters for the interaction energies of basket constituents, using physical analyses which we recently published. We also have begun to use FCM to examine processes occurring at the surfaces of intact cells, aimed, in part, at probing kinetic elements of ligand-induced receptor sequestration and receptor-mediated endocytosis. To infer possible triggering mechanisms for endocytosis and related intracellular membrane vesicle-forming processes, we have carried out mathematical and computational analysis of complex pathways of phosphatidylinositide biosynthesis and metabolism, showing that coupling of enzymatic activity to vesicle curvature can introduce nonlinearities and feedback loops leading to pseudo-bistability and triggering. In yet other studies we have used small angle neutron scattering (SANS) and ultra small angle x-ray scattering (USAXS) to study the supramolecular structure of tubulin polymers in solution, and have examined how such structure changes as a function of solvent composition. Finally, in cooperation with investigators from other laboratories, our staff has studied in vivo subcellular localization of tumor-suppressor protein p53 and the effects of antimitotic drugs on its association with, and transport along, microtubules.